Cytosolic isocitrate dehydrogenase, also known as isocitrate dehydrogenanse 1 (IDH1), is an enzyme that catalyzes the oxidative carboxylation of isocitrate to produce CO2 and α-ketoglutarate (αKG). Somatic mutations in IDH1 have been identified in colorectal cancers, gliomas (e.g., grade II-III astrocytomas, oligodendrogliomas and glioblastomas), patients with acute myeloid leukemia (AML), prostate, B-acute lymphoblastic leukemia, and a variety of other malignanices with lower frequencies (Sjöblom et. al., 2006, Science 314:268-274; Parsons et al., 2008, Science 321:1807-1812; Bleeker et al., 2009, Hum. Mutat. 30:7-11; Yan et al., 2009, N. Engl. J. Med. 360:765-773; Yan et al., 2009, N. Engl. J. Med. 360:765-773; Mardis et al., 2009, N. Engl. J. Med. 361:1058-1066). Importantly, gliomas and AML are two of the most malignant types of tumors, having the worst prognoses.
The IDH1 mutation found within these malignancies is a heterozygous mutation affecting Arg132, an amino acid that forms part of a catalytic arginine triad (along with Arg100 and Arg 109) involved in binding to isocitrate. Due the location of the mutation, it was originally hypothesized that the mutated protein would lack enzymatic activity; however, it was later discovered that rather than being catalytically inactive, the mutant form possesses the novel enzymatic activity of reducing αKG to 2-hydroxyglutarate (2HG) (Dang et al., 2009 Nature 462:739-744). The arginine at amino acid position 132 of IDH1 has been found to be mutated to histidine (R132H), cysteine (R132C), glycine (R132G), serine (R132S), leucine (R132L) and valine (R132V) (Yen et al., 2010, Oncogene 29:6409-6417).
PCT International patent application published as WO 2010/105243 (Dang et al.) describes methods for treating patients with cell proliferation-related disorders characterized by the presence of a mutant isocitrate dehydrogenase by administering a nucleic acid based inhibitor (e.g., siRNA) that targets mRNA encoding mutant IDH1 proteins that demonstrate 2HG neoactivity.
siRNA having specific sense and antisense sequences are provided in the application; however, the disclosure does not show either the down-regulation of IDH1 gene expression and/or decrease in IDH protein levels or the result of such down-regulation in any functional assays.
There remains a need for molecules that inhibit IDH1 and mutant forms thereof. Alteration of gene expression, specifically IDH1 and mutant IDH1 gene expression, through RNA interference (hereinafter “RNAi”) is one approach for meeting this need. RNAi is induced by short single-stranded RNA (“ssRNA”) or double-stranded RNA (“dsRNA”) molecules. The short dsRNA molecules, called “short interfering nucleic acids (“siNA”)” or “short interfering RNA” or “siRNA” or “RNAi inhibitors” silence the expression of messenger RNAs (“mRNAs”) that share sequence homology to the siNA. This can occur via cleavage of the mRNA mediated by an endonuclease complex containing a siNA, commonly referred to as an RNA-induced silencing complex (RISC). Cleavage of the target RNA typically takes place in the middle of the region complementary to the guide sequence of the siNA duplex (Elbashir et al., 2001, Genes Dev., 15:188). In addition, RNA interference can also involve small RNA (e.g., micro-RNA or miRNA) mediated gene silencing, presumably through cellular mechanisms that either inhibit translation or that regulate chromatin structure and thereby prevent transcription of target gene sequences (see for example Alishire, 2002, Science, 297:1818-1819; Volpe et al., 2002, Science, 297:1833-1837; Jenuwein, 2002, Science, 297:2215-2218; and Hall et al., 2002, Science, 297:2232-2237). Despite significant advances in the field of RNAi, there remains a need for agents that can inhibit IDH1 and mutant IDH1 gene expression and that can treat disease associated with mutant IDH1 expression, such as cancer.